title: Lactosylceramide interacts with and activates cytosolic phospholipase A2α.
abstract: Lactosylceramide (LacCer) is a member of the glycosphingolipid family and is known to be a bioactive lipid in various cell physiological processes. However, the direct targets of LacCer and cellular events mediated by LacCer are largely unknown. In this study, we examined the effect of LacCer on the release of arachidonic acid (AA) and the activity of cytosolic phospholipase A2α (cPLA2α). In CHO-W11A cells, treatment with 1-phenyl-2-palmitoylamino-3-morpholino-1-propanol (PPMP), an inhibitor of glucosylceramide synthase, reduced the glycosphingolipid level, and the release of AA induced by A23187 or platelet-activating factor was inhibited. The addition of LacCer reversed the PPMP effect on the stimulus-induced AA release. Exogenous LacCer stimulated the release of AA, which was decreased by treatment with an inhibitor of cPLA2α or silencing of the enzyme. Treatment of CHO-W11A cells with LacCer induced the translocation of full-length cPLA2α and its C2 domain from the cytosol to the Golgi apparatus. LacCer also induced the translocation of the D43N mutant of cPLA2α. Treatment of L929 cells with TNF-α induced LacCer generation and mediated the translocation of cPLA2α and AA release, which was attenuated by treatment with PPMP. In vitro studies were then conducted to test whether LacCer interacts directly with cPLA2α. Phosphatidylcholine vesicles containing LacCer increased cPLA2α activity. LacCer bound to cPLA2α and its C2 domain in a Ca(2+)-independent manner. Thus, we propose that LacCer is a direct activator of cPLA2α.
authors.affiliation: Laboratory of Chemical Pharmacology, Graduate School of Pharmaceutical Sciences, Chiba University, Chuo-ku, Chiba 260-8675, Japan. nakahiro@faculty.chiba-u.jp
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Nakamura
authors.forename: Yuta
authors.initials: Y
authors.lastname: Moriyama
authors.forename: Tomohiko
authors.initials: T
authors.lastname: Makiyama
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Emori
authors.forename: Hisahiro
authors.initials: H
authors.lastname: Yamashita
authors.forename: Risa
authors.initials: R
authors.lastname: Yamazaki
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Murayama
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000235:genetics / Q000378:metabolism*; D016718:Arachidonic Acid / Q000235:genetics / Q000378:metabolism; D016466:CHO Cells; D000001:Calcimycin / Q000494:pharmacology; D061207:Calcium Ionophores / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D020536:Enzyme Activators / Q000378:metabolism*; D005964:Glucosyltransferases / Q000235:genetics / Q000378:metabolism; D006056:Golgi Apparatus / Q000235:genetics / Q000378:metabolism*; D054513:Group IV Phospholipases A2 / Q000235:genetics / Q000378:metabolism*; D006168:Guinea Pigs; D006801:Humans; D007790:Lactosylceramides / Q000235:genetics / Q000378:metabolism*; D008614:Meperidine / Q000031:analogs & derivatives / Q000494:pharmacology; D051379:Mice; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: D015703:Antigens, CD; D061207:Calcium Ionophores; D020536:Enzyme Activators; D007790:Lactosylceramides; D014409:Tumor Necrosis Factor-alpha; C026733:4-propionyloxy-4-phenyl-N-methylpiperidine; D016718:Arachidonic Acid; D000001:Calcimycin; C009744:CDw17 antigen; D008614:Meperidine; D005964:Glucosyltransferases; C023246:ceramide glucosyltransferase; D054513:Group IV Phospholipases A2; C516389:PLA2G4A protein, human
keywords: Arachidonic acid; Glycosphingolipid; Phospholipase A; Sphingolipid; Tumor Necrosis Factor (TNF)